N-terminally-truncated and modified amyloid-beta (A) peptides are abundant in cerebral amyloid deposits in Alzheimer's disease (AD). Pyroglutamate A is generated upon N-terminal truncation of A followed by cyclization by glutaminyl cyclase to convert glutamic acid at residues 3 and 11 to pyroglutamate (ApE3 and ApE11). Both forms aggregate quickly, resist degradation, and are neurotoxic. It is unclear if either is present in initial A deposition in plaques and blood vessels (i.e., acting as a seed for further deposition) or if they are modified later. However, Alzheimer's disease progression appears to correlate with the presence of A pE peptide aggregates in brain. We hypothesize that pyroglutamate A acts as a seed for A deposition and accelerates inflammation, neurodegeneration and cognitive decline; therefore, targeted removal of this toxic species by immunotherapy will reduce A deposition, inflammation and neuritic dystrophy, and protect against cognitive impairment without disturbing non-pathogenic A. We propose 4 Specific Aims. Aim 1: We will determine if intrahippocampal or intraperitoneal injections of ApE-containing mouse brain extracts enhance A deposition, inflammation, neurodegeneration and cognitive decline in APP/PS1dE9 transgenic mice with aging in vivo. Aim 2: We will determine if early removal of pyroGlu A prevents general A plaque deposition and neuritic changes, and protects against cognitive decline by passively immunizing (i.p.) male APP/PS1dE9 tg mice with an anti-AN3pE mAb (07/1), an anti-ApE11 mAb, a general A mAb (3A1), or PBS weekly from 4-12 mo of age, starting prior to plaque onset. Outcome measures: behavioral, biochemical, and neuropathological analyses. Aim 3: We will determine if removal of pyroGlu A in late stage AD reduces total A and neurodegeneration and improves cognitive deficits by passively immunizing (i.p.) female APP/PS1dE9 tg mice weekly from 12-16 mo of age, starting well after plaque onset. Antibodies and outcome measures are the same as in Aim 2. Aim 4: We will examine the immune response of microglia to pyroGlu A mAbs in acute in vivo studies and in primary microglial cultures in vitro. Our collaborators at Probiodrug AG (Germany) will kindly provide us with 2 high-affinity, highly selective pyroGlu A mAbs (anti- A pE3 and anti-ApE11), synthetic ApE peptides, and brain extracts from their transgenic mouse models that accumulate pyroGlu-3 A peptides. Our collaborators at the CND have generously provided the 3A1 general A mAb hybridoma as a control. Importantly, my lab initiated this collaboration and has many years of experience investigating pyroGlu A deposition, inflammation, and A immunotherapy. The overall goal of our study is to determine whether pyroglutamate A proteins (ApE3 and ApE11) are therapeutic targets for Alzheimer's disease and, whether clearance by immunotherapy specific for either pyroGlu A species would be efficacious to prevent and/or treat AD. PUBLIC HEALTH RELEVANCE: Pyroglutamate A is a modified, truncated form of amyloid-beta (A) that is deposited into plaques and blood vessels in Alzheimer's disease brains. Pyroglutamate A (starting at amino acids 3 and 11) is resistant to degradation and is highly toxic in the brain. We will test whether these modified forms of A propagate amyloid deposition, neurodegeneration, and cognitive impairment and, whether removal of them can prevent or treat Alzheimer's disease in a mouse model.